Related Experiment Video
Updated: May 11, 2026
In Vitro Assay to Evaluate the Impact of Immunoregulatory Pathways on HIV-specific CD4 T Cell Effector Function
Published on: October 15, 2013
Integrated Chronic In Vivo and In Vitro Screens Uncover NFIL3 as a Driver of T-cell Dysfunction
Nayan Jain1, Yuzhe Shi1, Celina May2,3
1Columbia Initiative in Cell Engineering and Therapy (CICET), Vagelos College of Physicians and Surgeons, Columbia University Irving Medical Center, Columbia University, New York, New York.
Abstract:
Chimeric antigen receptor (CAR) therapy has transformed the treatment landscape for hematologic malignancies, but its efficacy in solid tumors is limited, owing in part to insufficient functional persistence of the engineered T cells. To elucidate the basis for their functional decline, we conducted integrated chronic in vivo and in vitro screens of 400 transcription factors, which revealed NFIL3 as a driver of CAR T-cell dysfunction. Genetic disruption of NFIL3 in CAR T cells sustains their expansion and increases cytokine production, overall restraining terminal differentiation. Loss of NFIL3 enhances CAR T-cell efficacy, improving tumor control and prolonging survival in xenograft and syngeneic mouse tumor models across different CAR designs. Under chronic stimulation, disruption of NFIL3 establishes a transcriptional state predictive of favorable clinical outcomes. Our findings underscore the power of comprehensive in vivo genetic screens integrated with multiparameter in vitro assessment and identify NFIL3 as a novel therapeutic target to enhance cancer immunotherapy.
Significance:
This study presents a two-step screening framework, integrating an in vivo pooled guide RNA screen with a multiparameter, in vitro arrayed screen. NFIL3 emerged as the top candidate, and its disruption enhanced CAR T-cell antitumor efficacy in both hematologic malignancies and solid tumors across diverse CAR architectures.
Insights
NFIL3 drives dysfunction in CAR T cells, limiting their effectiveness against solid tumors. Disrupting NFIL3 enhances CAR T cell persistence and anti-tumor activity, offering a new therapeutic target for cancer immunotherapy.
Area of Science:
- Immunology
- Molecular Biology
- Oncology
Background:
- Chimeric antigen receptor (CAR) T cell therapy shows promise in hematological cancers but faces challenges in solid tumors due to limited T cell persistence.
- Understanding the molecular mechanisms behind CAR T cell dysfunction is crucial for improving their efficacy.
Purpose of the Study:
- To identify key regulators of CAR T cell dysfunction during chronic stimulation.
- To evaluate the therapeutic potential of targeting identified regulators to enhance CAR T cell function in solid tumors.
Main Methods:
- Conducted integrated chronic in vivo and in vitro screens of 400 transcription factors.
- Utilized genetic disruption of identified factors in CAR T cells.
- Assessed CAR T cell expansion, cytokine production, differentiation, and anti-tumor efficacy in mouse models.
Main Results:
- Identified NFIL3 as a critical driver of CAR T cell dysfunction.
- Genetic disruption of NFIL3 sustained CAR T cell expansion and cytokine production while restraining terminal differentiation.
- Loss of NFIL3 enhanced CAR T cell efficacy, improving tumor control and survival in preclinical models.
- NFIL3 disruption induced a transcriptional state associated with favorable clinical outcomes.
Conclusions:
- NFIL3 is a key determinant of CAR T cell functional decline in the tumor microenvironment.
- Targeting NFIL3 represents a promising strategy to enhance the efficacy of CAR T cell therapy for solid tumors.
- Integrated in vivo and in vitro genetic screening is a powerful approach for discovering therapeutic targets in cancer immunotherapy.
